Figure 1
In vitro characterization of PR8-H5-NS1(73)H5 virus. (A) Schematic representation of promoters and coding sequences of pHW-NS1(73)-H5 HA1-NEP plasmid used to generate recombinant influenza virus. (B) Growth kinetics of PR8-H5-NS1(73)H5 and PR8-2344H5 in ECEs: 104 EID50 per virus was inoculated into ECEs. At 4, 8, 12, 24, 36, and 48 h p.i., allantoic fluids were collected and titrated in ECEs. Shown is mean with standard error for each data point. (C) Genetic stability of PR8-H5-NS1(73)H5 in chicken embryos, by RT-PCR. 1) pHW plasmid encoding NS1(73)-H5 HA1-NEP genes; 2) Passage 1 of PR8-H5-NS1(73)H5; 3) Passage 3 of PR8-H5-NS1(73)H5; 4); Passage 5 of PR8-H5-NS1(73)H5; 5) Passage 10 of PR8-H5-NS1(73)H5. (D) ECEs infected with PR8-2344H5, PR8-2321H5, or PR8-H5-NS1(73)H5 virus with allantoic fluids purified by ultracentrifugation through a 20% sucrose cushion. Proteins were visualized by western blot using anti-H5 against a different clade or anti-NS1 monoclonal antibody.
